Lecture 29 Regulation of Gene Expression

Question 1

Linkage analysis

Answer 1

• could take years
• looked at X-chromosomes and autosomes because they undergo recombination
• Y chromosome does not undergo recombination

Question 2

Exome sequencing

Answer 2

• allows us to identify causal mutations in those cases where a large family suitable for linkage analysis is not available
• only takes a couple of individuals
• relatively inexpensive (few thousand dollars)

Question 3

Gene nomenclature

Answer 3

• all letters in the gene are italicized
• human gene names are designated with capitalized and italicized letters
• Mouse genes are designated with the 1st letter capitalized and all letters italicized
• LRP5 vs Lrp5(mouse)

Question 4

DNA sequencing

Answer 4

• genome sequencing
• exons
• mRNA (transcriptomic sequencing or RNA-seq)
• Epigenomics

Question 5

Genome sequencing

Answer 5

• every base in the human genome or mouse

- putting it in correct order

Question 6

Exomic

Answer 6

• only on exons or protein coding information

Question 7

mRNA sequencing

Answer 7

• messenger RNA population in different tissues

- which genes are being expressed at that time of extraction

Question 8

Epigenomics

Answer 8

• DNA methylation
• ChIP sequencing
• Ribosome profiling

Question 9

cost of DNA sequencing

Answer 9

• becoming more and more affordable

Question 10

Important questions in regulation of gene expression?

Answer 10

• how is the DNA sequence information read by the cell

- What is the relationship between the start site of transcription and the location of the control sequences?

Question 11

Whats a gene?

Answer 11

• A gene is defined as a segment of DNA that is transcribed into RNA and its associated transcriptional control regions
• Not just the information that gives rise to the protein, its the exons, introns, and untranslated regions and transcriptional control regions

Question 12

Levels of gene expression control

Answer 12

• gene requirements

- gene specific requirements

Question 13

Gene requirements

Answer 13

• transcriptional machinery (must be present for transcription to occur)

Question 14

Gene specific requirements

Answer 14

• intracellular hormone- receptor complexes
  (steroids (estrogen, testosterone, Vit D3 etc.) and their cognate receptors)
• Intracellular second messengers
  (peptide hormones that bind to cell membrane receptors)
• Gene specific (class specific) transcription factors
• Gene regulatory sequences

Question 15

DNA Binding protein Motifs

Answer 15

• Helix-loop- helix
• Helix- turn-helix
• Leucine Zipper

Question 16

Helix-loop-helix

Answer 16

• common in transcription factors and consists of alpha helices bound by a looping stretch of amino acids

Question 17

Zinc Finger Motifs

Answer 17

• multiple types but all have a bound zinc metal ion associated with the protein. Some have an alpha helix plus beta sheet structure, others have a more helix turn helix structure. Can be monomers or dimers

Question 18

Helix-turn-helix

Answer 18

• DNA binding motif consisting of two alpha helices joined by a short stretch of amino acids

Question 19

Leucine zipper

Answer 19

• two proteins each with an alpha helix form a Y shaped coiled-coil structure. Results in both protein and DNA binding

Question 20

Important DNA sequences in Pol II transcribed genes

Answer 20

• TATA box
• INR
• DPE
• BRE

Question 21

TATA box

Answer 21

• located ~ 25-30 bp 5’ of the start site of transcripts

- binds to the TBP subunit of TFIID

Question 22

INR

Answer 22

• contains start site of transcription for many RNA Pol II genes, binds TFIID

Question 23

DPE

Answer 23

• located at +30 (3’) of the start site

- binds TFIID

Question 24

BRE:

Answer 24

• located at -35 (5’) of the start site,

- Binds TFIIB

Question 25

Is DNA a rigid linear structure

Answer 25

• no DNA can loop back on itself

- why regulatory proteins can be far away from the start site (but close in 3D)

Question 26

Summary of activation of Transcription at a specific promoter

Answer 26

1. Gene activator protein binds to chromatin
2. Chromatin Remodeling
3. Covalent histone modification
4. Additional activator proteins bound to gene regulatory region
5. Assembly of pre-initiation complex at the promoter
6. Transcription initiation

Question 27

Multiple levels of control of Eukaryotic gene expression, why is it neccessary?

Answer 27

• necessary so gene is only expressed when you need it and protein is only made when you need it

Question 28

What are the different methods of regulating gene expression/transcription?

Answer 28

• Transcriptional control
• RNA processing control
• RNA transport and localization control
• Translation control
• mRNA degradation control (miRNA)
• Protein activity control

Question 29

What 5 families/pathways work together to turn a fertilized egg into a human?

Answer 29

1. Receptor Tyrosine Kinase
2. TGFB superfamily
3. Wnt
4. Hedgehog
5. Notch

Question 30

Why are B-cattenin levels low in the cell?

Answer 30

• because they signal many growth pathways

Question 31

How are B cat levels low?

Answer 31

• degredation complex

Question 32

What happens when wnt is present?

Answer 32

• B cat is no longer degreaded and it can translocate into the nucleus and binds to TCF (which is bound to the DNA)

Question 33

Mutation in wnt/B-catenin pathway can lead to what?

Answer 33

-cancer

Question 34

Dkk1

Answer 34

• binds to Lrp5/6 which gets it degraded so wnt can function

Question 35

Sclerostin

Answer 35

-binds to Lrp5/6 which prevents wnt from binding

Question 36

Frizzled

Answer 36

-homologs which bind to wnt so it cant bind to the Lrp5/6

Question 37

Wise

Answer 37

• binds to Lrp6